To investigate the role of the monomeric guanosine triphosphatase (GTPase) Rho on migration of hematopoietic progenitor cells (HPC), we employed different clostridial toxins which inhibit the Rho family of GTPases. Pretreatment with C2I‐C3, a cell‐accessible C3 transferase fusion protein that targets Rho, increased chemokinetic migration of the factor‐dependent multipotent cell line Factor Dependent Cell Paterson with mixed lineage differentiation potential (FDCP‐mix) and of primary lineage marker‐depleted HPC in vitro. In contrast, treatment with lethal toxin (LT) from Clostridium sordellii, which predominantly inactivates Rac, and with toxin B from C. difficile, which inactivates Rho, Rac and Cdc42, decreased in vitro migration. When HPC pretreated with LT or toxin B were transplanted into mice, homing to the bone marrow was impaired, whereas C2I‐C3 treatment did not alter HPC homing. However, in a competitive hematopoietic repopulation experiment in C57BL/6 mice, pretreatment of bone marrow cells with any of the inhibitors, including the Rho inhibitor C2I‐C3, resulted in suppressed donor‐type hematopoiesis. Our data indicate that whereas Rac supports HPC cell cycling, migration, short‐term homing and hematopoietic regeneration, Rho coordinates down‐regulation of HPC migration and is required for hematopoietic regeneration.
The oncogene E2a-Pbx1 is formed by the t(1;19) translocation, which joins the N-terminal transactivation domain of E2a with the C-terminal homeodomain of PBX1. The goal of this work was to elucidate the mechanisms by which E2a-Pbx1 can lead to deregulated target gene expression. For reporter constructs it was shown that E2a-Pbx1 can activate transcription through homodimer elements (TGATTGAT) or through heterodimer elements with Hox proteins (e.g. TGATTAAT). We show a novel mechanism by which E2a-Pbx1 activates transcription of EF-9 using a promoter in intron 1 of the EF-9 gene, resulting in an aminoterminal truncated transcript. Our results indicate that the LDFS motif of E2a is essential for the transactivation of EF-9, but dispensable for transactivation of fibroblast growth factor 15. The E2a LDFS motif was also essential for proliferation of NIH3T3 fibroblasts but was dispensable for the E2a-Pbx1-induced differentiation arrest of myeloid progenitors.
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